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• Tâche 1: Identification of QTL, allelic variants of some relevant genes that have a significant effect on the grain contents in As, Cd, Ni, Pb and to examine in parallel their effects on grain Fe, Zn, P and phytates and on yield.

A collection of 220 bread winter wheat  and 200 durum wheat genotypes will be characterized for their contents in  As, Cd, Ni, Pb, Fe, Zn and P. Valuable QTLs associated to an optimized grain trace element (TE) content. Genome wide Association studies will be conducted to identify QTL associated to a low content in contaminants with no or minimum impacts on yield and on the contents in Fe and Zn.  For the relevant regions, Kaspar markers will be designed and validated for screening new genotypes (to identify interesting cultivars for WP4 trials) and for breeding and varietal characterization beyond the project.

Besides genes under GWAS confidence intervals (se above), the diversity of two gene families will be investigated: NAM-1 (No Apical Meristem) genes that facilitate nutrient remobilization from vegetative parts into grains and genes involved in the root uptake and partitioning of TEs (genes coding for ZIP and HMA proteins). The effects of allelic variants on grain content in As, Cd, Ni, Pb, Fe, Zn and P and on yield will be examined.

• Tâche 2: Identification of some relevant ecophysiological plant traits associated to a low grain content in As, Cd, Ni, Pb without important negative effects grain Fe, Zn, P and phytates and on yield.

From WP1, a subset of 40 bread wheat and 20 durum wheat cultivars maximizing the variability in grain TE contents and associated QTLs and alleles will be grown in field trials at two locations (different pedoclimatic and TE bioavailability conditions) to examine before anthesis and at physiological maturity the possible correlations between grain TE contents, yield and the traits assumed to be linked to grain TE (see figure): biomass partitioning, plant architecture, proportion of tissues involved in TE transport and storage, plant functioning including TE mobilization and uptake in the rhizosphere, phenology and nutrient remobilization (Yan et al., 2018), which will be followed by non-destructive portable X-ray fluorescence. Traits related to roots being hardly measurable in the field, they will be determined on one-month-old seedlings grown in rhizoboxes.

• Tâche 3: Mechanistic study of TE partitioning to and within the spike

- WP 3.1: Allocation of TEs to spike grains depending on spike structure and functioning. The relative importance of (xylem+apoplast) versus phloem pathways of TEs to grains will be studied. Spike, spikelet and grain transpiration will be manipulated by awn removal and by modifying the evapotranspiration demand (VPD) in the spike environment. Phloem transport will be reduced by steam girdling of the peduncle. The competition between spikelets and between grains within a spikelet will be studied by removing spikelets and grains at different ranks of the spike. These analyses will be performed in controlled conditions for 2 genotypes of durum and bread wheat having contrasted spike structure and TE contents.

- WP 3.2: Allocation of TEs to grains depending on TE trafficking at the peduncle node. The peduncle node being central for TE trafficking to grains, the relative abundance of TE at this location will be examined at early and late grain filling stages for cultivars of WP 3.1 by synchrotron micro X-ray fluorescence (µXRF). The ligands to which TEs are associated in the various parts of the node will be investigated by micro X-ray Absorption Near Edge Structure spectroscopy (µXanes). The distribution of contaminants and micronutrients in grains will be quantitatively established in parallel and compared to their distribution in nodes for the selected genotypes.

• Tâche 4: Test of markers and traits in contrasted environmental conditions

From WPs 1, 2 and 3, 15 bread wheat and 10 durum wheat cultivars having the extreme combinations of interesting traits, QTLs, alleles and grain TE contents will be tested in the field, for 2 years, at 4 experimental stations (INRAE, Arvalis, Florimond-Desprez and RAGT) with contrasted pedoclimatic conditions and soil TE bioavailability. We will test the stability of the association between the grain TE contents and the genetic markers+relevant phenotypic traits identified in WPs 1, 2 and 3.

• Tâche 5: Costs / benefits of markers and traits and outcomes for breeding /growing safe wheats

From the field data of WP’s 2 and 4, the grain TE contents will be modelled from the selected traits and markers in order to derive a positive or negative score for each of the latter, reflecting their effects on the grain contaminant and micronutrient contents and on yield. Theses scores will rank the contribution of traits and genetic markers in the perspective of obtaining wheat grains with optimized TE contents. This will help wheat breeders to predict the expected trade-off they might have to accept in their breeding strategy. The expertise of Florimond-Desprez and of RAGT will help to decide which markers and traits are practically useful in a breeding strategy. The WP5 will also establish the scoring for existing cultivars, and thanks to the dissemination by Arvalis, this will help farmers to minimize the risk of non-conformity of their harvests.